Study of the effect of protein synthesis inhibitors on growing <i>Escherichia coli</i> bacteria using electrochemical sensors

Abstract

Background. The study of the mechanisms of action of antibiotics requires the integrated use of traditional microbiological and physicochemical methods. The aim. To study the response of Escherichia coli bacteria to the action of four antibiotics, inhibitors of protein synthesis, using combined approach. Methods. Bacteria were grown under aerobic conditions on minimal M9 medium with glucose. Tetracycline, kanamycin, streptomycin and chloramphenicol have been tested. The effect of antibiotics on survival (CFU) and growth rate was determined. Respiratory activity, sulfide production, extracellular potassium, as well as pH and Eh of the medium were measured using electrochemical sensors directly in a growing culture in the “real time”, membrane potential was measured using a DiBAC dye and a Leica DM2000 fluorescent microscope. Results. The tested antibiotics were divided into two groups according to their properties. Tetracycline and chloramphenicol showed a pronounced bacteriostatic effect, growth inhibition began immediately after the addition of antibiotics and occurred at a high rate. Both antibiotics inhibited respiration, stimulated sulfide production and an Eh jump. Bacteria treated with tetracycline and chloramphenicol retained the ability to maintain membrane potential and intracellular potassium better. Inhibition of respiration led to a decrease in glucose catabolism, as evidenced by a lower rate of acidification of the medium compared to the control. Growth inhibition with streptomycin and kanamycin was initiated with a 30-minute delay. Both antibiotics showed a bactericidal effect, did not stimulate sulfide production and Eh jump, did not inhibit respiration, but caused a drop in membrane potential and intracellular potassium. High respiratory activity promoted glucose catabolism, as evidenced by the rapid acidification of the medium. Of interest is the detection of kanamycin-induced sulfide production during E. coli growth on MOPS medium. Conclusion. An analysis of the data obtained indicates that the use of electrochemical sensors in combination with traditional methods is a promising approach to studying the mechanisms of action of antibiotics.